RESUMO
Mitochondria shape intracellular Ca2+ signaling through the concerted activity of Ca2+ uptake via mitochondrial calcium uniporters and efflux by Na+ /Ca2+ exchangers (NCLX). Here, we describe a novel relationship among NCLX, intracellular Ca2+ , and autophagic activity. Conditions that stimulate autophagy in vivo and in vitro, such as caloric restriction and nutrient deprivation, upregulate NCLX expression in hepatic tissue and cells. Conversely, knockdown of NCLX impairs basal and starvation-induced autophagy. Similarly, acute inhibition of NCLX activity by CGP 37157 affects bulk and endoplasmic reticulum autophagy (ER-phagy) without significant impacts on mitophagy. Mechanistically, CGP 37157 inhibited the formation of FIP200 puncta and downstream autophagosome biogenesis. Inhibition of NCLX caused decreased cytosolic Ca2+ levels, and intracellular Ca2+ chelation similarly suppressed autophagy. Furthermore, chelation did not exhibit an additive effect on NCLX inhibition of autophagy, demonstrating that mitochondrial Ca2+ efflux regulates autophagy through the modulation of Ca2+ signaling. Collectively, our results show that the mitochondrial Ca2+ extrusion pathway through NCLX is an important regulatory node linking nutrient restriction and autophagy regulation.
Assuntos
Sinalização do Cálcio , Cálcio , Clonazepam/análogos & derivados , Tiazepinas , Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Trocador de Sódio e Cálcio , Mitocôndrias/metabolismo , Autofagia , Sódio/metabolismoRESUMO
Significance: The systematic investigation of oxidative modification of proteins by reactive oxygen species started in 1980. Later, it was shown that reactive nitrogen species could also modify proteins. Some protein oxidative modifications promote loss of protein function, cleavage or aggregation, and some result in proteo-toxicity and cellular homeostasis disruption. Recent Advances: Previously, protein oxidation was associated exclusively to damage. However, not all oxidative modifications are necessarily associated with damage, as with Met and Cys protein residue oxidation. In these cases, redox state changes can alter protein structure, catalytic function, and signaling processes in response to metabolic and/or environmental alterations. This review aims to integrate the present knowledge on redox modifications of proteins with their fate and role in redox signaling and human pathological conditions. Critical Issues: It is hypothesized that protein oxidation participates in the development and progression of many pathological conditions. However, no quantitative data have been correlated with specific oxidized proteins or the progression or severity of pathological conditions. Hence, the comprehension of the mechanisms underlying these modifications, their importance in human pathologies, and the fate of the modified proteins is of clinical relevance. Future Directions: We discuss new tools to cope with protein oxidation and suggest new approaches for integrating knowledge about protein oxidation and redox processes with human pathophysiological conditions. Antioxid. Redox Signal. 35, 1016-1080.
Assuntos
Proteínas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Humanos , Oxirredução , Transdução de SinaisRESUMO
Significance: The systematic investigation of oxidative modification of proteins by reactive oxygen species started in 1980. Later, it was shown that reactive nitrogen species could also modify proteins. Some protein oxidative modifications promote loss of protein function, cleavage or aggregation, and some result in proteotoxicity and cellular homeostasis disruption. However, not all oxidative modifications are necessarily associated with damage, as with Met and Cys protein residue oxidation. In these cases, redox state changes can alter protein structure, catalytic function, signaling processes in response to metabolic and/or environmental alterations. This review aims to integrate the present knowledge on redox modifications of proteins with their fate and role in redox signaling and human pathological conditions. Critical issues: It is hypothesized that protein oxidation participates in the development and progression of many pathological conditions. However, no quantitative data has been correlated with specific oxidized proteins or the progression or severity of pathological conditions. Hence, the comprehension of the mechanisms underlying these modifications, their importance in human pathologies and, the fate of the modified proteins is of clinical relevance. Future directions: We discuss new tools to cope with protein oxidation and suggest new approaches for integrating knowledge about protein oxidation and redox processes with human pathophysiological conditions.
RESUMO
Aging is often accompanied by a decline in mitochondrial mass and function in different tissues. Additionally, cell resistance to stress is frequently found to be prevented by higher mitochondrial respiratory capacity. These correlations strongly suggest mitochondria are key players in aging and senescence, acting by regulating energy homeostasis, redox balance and signalling pathways central in these processes. However, mitochondria display a wide array of functions and signalling properties, and the roles of these different characteristics are still widely unexplored. Furthermore, differences in mitochondrial properties and responses between tissues and cell types, and how these affect whole body metabolism are also still poorly understood. This review uncovers aspects of mitochondrial biology that have an impact upon aging in model organisms and selected mammalian cells and tissues.
Assuntos
Envelhecimento/fisiologia , Mitocôndrias/metabolismo , Células-Tronco Adultas/metabolismo , Animais , Encéfalo/metabolismo , Caenorhabditis elegans/metabolismo , Metabolismo Energético/fisiologia , Humanos , Modelos Biológicos , Leveduras/metabolismoRESUMO
Calorie restriction (CR) is an intervention known to extend the lifespan of a wide variety of organisms. In S. cerevisiae, chronological lifespan is prolonged by decreasing glucose availability in the culture media, a model for CR. The mechanism has been proposed to involve an increase in the oxidative (versus fermentative) metabolism of glucose. Here, we measured wild-type and respiratory incompetent (ρ(0)) S. cerevisiae biomass formation, pH, oxygen and glucose consumption, and the evolution of ethanol, glycerol, acetate, pyruvate and succinate levels during the course of 28 days of chronological aging, aiming to identify metabolic changes responsible for the effects of CR. The concomitant and quantitative measurements allowed for calculations of conversion factors between different pairs of substrates and products, maximum specific substrate consumption and product formation rates and maximum specific growth rates. Interestingly, we found that the limitation of glucose availability in CR S. cerevisiae cultures hysteretically increases oxygen consumption rates many hours after the complete exhaustion of glucose from the media. Surprisingly, glucose-to-ethanol conversion and cellular growth supported by glucose were not quantitatively altered by CR. Instead, we found that CR primed the cells for earlier, faster and more efficient metabolism of respiratory substrates, especially ethanol. Since lifespan-enhancing effects of CR are absent in respiratory incompetent ρ(0) cells, we propose that the hysteretic effect of glucose limitation on oxidative metabolism is central toward chronological lifespan extension by CR in this yeast.
Assuntos
Saccharomyces cerevisiae/metabolismo , Biomassa , Restrição Calórica , Respiração Celular , Sobrevivência Celular , Meios de Cultura/química , Metabolismo Energético , Glucose/metabolismo , Concentração de Íons de Hidrogênio , Mitocôndrias/metabolismo , Oxirredução , Oxigênio/metabolismo , Saccharomyces cerevisiae/citologia , Fatores de TempoRESUMO
Knowledge of location and intracellular subcompartmentalization is essential for the understanding of redox processes, because oxidants, owing to their reactive nature, must be generated close to the molecules modified in both signaling and damaging processes. Here we discuss known redox characteristics of various mitochondrial microenvironments. Points covered are the locations of mitochondrial oxidant generation, characteristics of antioxidant systems in various mitochondrial compartments, and diffusion characteristics of oxidants in mitochondria. We also review techniques used to measure redox state in mitochondrial subcompartments, antioxidants targeted to mitochondrial subcompartments, and methodological concerns that must be addressed when using these tools.
Assuntos
Compartimento Celular , Mitocôndrias/fisiologia , Estresse Oxidativo , Animais , Apoptose , Humanos , Oxirredução , Espécies Reativas de Oxigênio/metabolismo , Transdução de SinaisRESUMO
Calorie restriction (CR) enhances animal life span and prevents age-related diseases, including neurological decline. Recent evidence suggests that a mechanism involved in CR-induced life-span extension is NO(â¢)-stimulated mitochondrial biogenesis. We examine here the effects of CR on brain mitochondrial content. CR increased eNOS and nNOS and the content of mitochondrial proteins (cytochrome c oxidase, citrate synthase, and mitofusin) in the brain. Furthermore, we established an in vitro system to study the neurological effects of CR using serum extracted from animals on this diet. In cultured neurons, CR serum enhanced nNOS expression and increased levels of nitrite (a NO(â¢) product). CR serum also enhanced the levels of cytochrome c oxidase and increased citrate synthase activity and respiratory rates in neurons. CR serum effects were inhibited by L-NAME and mimicked by the NO(â¢) donor SNAP. Furthermore, both CR sera and SNAP were capable of improving neuronal survival. Overall, our results indicate that CR increases mitochondrial biogenesis in a NO(â¢)-mediated manner, resulting in enhanced reserve respiratory capacity and improved survival in neurons.
Assuntos
Encéfalo/metabolismo , Restrição Calórica , Radicais Livres/metabolismo , Mitocôndrias/metabolismo , Neurônios/citologia , Óxido Nítrico/metabolismo , Sistema Respiratório/metabolismo , Animais , Western Blotting , Encéfalo/citologia , Células Cultivadas , Citrato (si)-Sintase , Feminino , Camundongos , Neurônios/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , OxirreduçãoRESUMO
eNOS activation resulting in mitochondrial biogenesis is believed to play a central role in life span extension promoted by calorie restriction (CR). We investigated the mechanism of this activation by treating vascular cells with serum from CR rats and found increased Akt and eNOS phosphorylation, in addition to enhanced nitrite release. Inhibiting Akt phosphorylation or immunoprecipitating adiponectin (found in high quantities in CR serum) completely prevented the increment in nitrite release and eNOS activation. Overall, we demonstrate that adiponectin in the serum from CR animals increases NO⢠signaling by activating the insulin pathway. These results suggest this hormone may be a determinant regulator of the beneficial effects of CR.
Assuntos
Adiponectina/metabolismo , Restrição Calórica , Insulina/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Adiponectina/sangue , Animais , Células Cultivadas , Endotélio Vascular/citologia , Proteína Oncogênica v-akt/metabolismo , Fosforilação , Ratos , Transdução de SinaisRESUMO
Calorie restriction is a dietary intervention known to improve redox state, glucose tolerance, and animal life span. Other interventions have been adopted as study models for caloric restriction, including nonsupplemented food restriction and intermittent, every-other-day feedings. We compared the short- and long-term effects of these interventions to ad libitum protocols and found that, although all restricted diets decrease body weight, intermittent feeding did not decrease intra-abdominal adiposity. Short-term calorie restriction and intermittent feeding presented similar results relative to glucose tolerance. Surprisingly, long-term intermittent feeding promoted glucose intolerance, without a loss in insulin receptor phosphorylation. Intermittent feeding substantially increased insulin receptor nitration in both intra-abdominal adipose tissue and muscle, a modification associated with receptor inactivation. All restricted diets enhanced nitric oxide synthase levels in the insulin-responsive adipose tissue and skeletal muscle. However, whereas calorie restriction improved tissue redox state, food restriction and intermittent feedings did not. In fact, long-term intermittent feeding resulted in largely enhanced tissue release of oxidants. Overall, our results show that restricted diets are significantly different in their effects on glucose tolerance and redox state when adopted long-term. Furthermore, we show that intermittent feeding can lead to oxidative insulin receptor inactivation and glucose intolerance.
Assuntos
Restrição Calórica/métodos , Dieta Redutora/métodos , Gordura Intra-Abdominal/metabolismo , Músculo Esquelético/metabolismo , Obesidade/dietoterapia , Receptor de Insulina/metabolismo , Adiposidade , Animais , Western Blotting , Peso Corporal , Glucose/metabolismo , Intolerância à Glucose/metabolismo , Humanos , Insulina/metabolismo , Proteínas Substratos do Receptor de Insulina/análise , Proteínas Substratos do Receptor de Insulina/biossíntese , Masculino , Óxido Nítrico Sintase/análise , Óxido Nítrico Sintase/biossíntese , Nitrocompostos , Obesidade/metabolismo , Oxirredução , Ratos , Ratos Sprague-Dawley , Receptor de Insulina/antagonistas & inibidoresRESUMO
Mild mitochondrial uncoupling, or the reduction of the efficiency of energy conversion without compromising intracellular high energy phosphate levels, is a protective therapeutic strategy under many laboratory conditions. Here we discuss these conditions, which include both cell and animal models of ischemia reperfusion and complications associated with the metabolic syndrome. We also discuss drugs that promote mild mitochondrial uncoupling and naturally occurring mild mitochondrial uncoupling pathways involving free fatty acid cycling and K(+) transport.